Dealing with uncertain results from chromosomal microarray and exome sequencing in the prenatal setting: an international cross-sectional study with healthcare professionals.

OBJECTIVES: To conduct qualitative interviews with healthcare providers working in different countries to understand their experiences of dealing with uncertain results from prenatal chromosome microarray analysis (CMA) and exome sequencing (ES). METHODS: Semi-structured interviews with 31 healthcare providers who report or return prenatal CMA and/or ES results (clinicians, genetic counsellors and clinical scientists) in six countries with differing healthcare systems; Australia (4), Denmark (5), Netherlands (6), Singapore (4), Sweden (6) and United Kingdom (6). The topic guide explored the main sources of uncertainty and their management. RESULTS: There was variation in reporting practices both between and across countries for variants of uncertain significance (VUS), however, there was broad agreement on reporting practices for incidental findings. There was also variation in who decides what results are reported (clinical scientists or clinicians). Technical limitations and lack of knowledge (to classify variants and of prenatal phenotypes) were significant challenges, as were turnaround times and lack of guidelines. CONCLUSION: Health professionals around the globe are dealing with similar sources of uncertainty, but managing them in different ways, Continued dialogue with international colleagues on ways of managing uncertain results is important to compare and contrast the benefits and limitations of the different approaches. This article is protected by copyright. All rights reserved

SITC cancer immunotherapy resource document: a compass in the land of biomarker discovery.

Since the publication of the Society for Immunotherapy of Cancer\u27s (SITC) original cancer immunotherapy biomarkers resource document, there have been remarkable breakthroughs in cancer immunotherapy, in particular the development and approval of immune checkpoint inhibitors, engineered cellular therapies, and tumor vaccines to unleash antitumor immune activity. The most notable feature of these breakthroughs is the achievement of durable clinical responses in some patients, enabling long-term survival. These durable responses have been noted in tumor types that were not previously considered immunotherapy-sensitive, suggesting that all patients with cancer may have the potential to benefit from immunotherapy. However, a persistent challenge in the field is the fact that only a minority of patients respond to immunotherapy, especially those therapies that rely on endogenous immune activation such as checkpoint inhibitors and vaccination due to the complex and heterogeneous immune escape mechanisms which can develop in each patient. Therefore, the development of robust biomarkers for each immunotherapy strategy, enabling rational patient selection and the design of precise combination therapies, is key for the continued success and improvement of immunotherapy. In this document, we summarize and update established biomarkers, guidelines, and regulatory considerations for clinical immune biomarker development, discuss well-known and novel technologies for biomarker discovery and validation, and provide tools and resources that can be used by the biomarker research community to facilitate the continued development of immuno-oncology and aid in the goal of durable responses in all patients

Point-of-care diagnostics of childhood central nervous system infections, with a focus on usability in low-resource settings

Background: The inaccessibility of laboratory services sustains the high burden of paediatric infectious diseases, such as central nervous system (CNS) infections, in low-resource settings. New contextually fit and well-implemented point-of-care tests (POCTs) could relieve such a burden and narrow the diagnostic divide between rich and poor. Yet, current disengagement between product developers, end-users, and implementors of POCTs impedes their clinical use and utility in low-resource settings. Also, the lack of evidence gathered through field evaluations of many diagnostic instruments in low-resource settings raises questions of their clinical utility there. Objectives: The main aim of this thesis was to provide clinical and contextual guidance for developers of new POCTs for CNS infection diagnosis with high utility, especially in low-resource settings; and to implementors of POCT services towards their optimized clinical benefit. This was addressed through a multidisciplinary combination of qualitative, laboratory, and clinical studies. Methods: Qualitative focus group discussions were conducted with health care workers (HCW) in Mbarara, Uganda (Paper I), and in Stockholm, Sweden (Paper III). Discussions were audio recorded and transcribed verbatim. Qualitative content analysis with an inductive approach was pursued in for data analysis. Comparisons between the two settings were discussed. In Paper II, a vertical flow DNA microarray printed on paper was developed for the detection of Neisseria meningitidis – a major aetiology of paediatric bacterial CNS infection worldwide. The analytical performance of the microarray was laboratory evaluated on DNA extracted from the bacteria, through the detection of the ctrA gene sequence specific to N. meningitidis. In Paper IV, a commercially available polymerase chain reaction (PCR) instrument with the capability of multiplex single-sample cerebrospinal fluid (CSF) microbiology was prospectively field-evaluated for the diagnosis of paediatric CNS infection in Mbarara, Uganda. Clinical turnaround time (cTAT) was defined as time spent from lumbar puncture until reporting of microbiology analyses to clinicians. The PCR instrument’s influence on clinical and patient-centered outcomes (yield, cTAT, duration of hospitalization and antibiotic exposure, patient outcome) was compared to that of bacterial culture. Results: Fifty and 24 HCWs of different professions participated in the qualitative studies in Mbarara and Stockholm, respectively, expressing greater similarities than differences in perspectives of POCT use. POCTs were routinely used at both sites and credited for facilitating differential diagnostics and clinical decision-making. While the Ugandan setting with low laboratory accessibility was highly dependent on POCTs for sample analyses, the Swedish setting credited their use for having clinical and social value. Contrary to the described beneficial aspects, current POCTs were deemed contextually unfit in Mbarara, and their use to cause clinical distraction in Stockholm. Deficient implementation of POCT services was exposed in both places. Requests for ideal POCTs were aligned with those stipulated by the ‘ASSURED’ criteria of the World Health Organization. Specific POCTs for infectious diseases, including CNS infections, were requested. The laboratory study demonstrated an analytical sensitivity of 38 copies of ctrA per assay, with high specificity. The clinical study enrolled 212 children aged 0-12 years who were suspected of having CNS infection, with 193 of them being evaluated using the commercially available PCR instrument. A vast majority of children had been pre-administered antibiotics prior to lumbar puncture. Bacterial yield for the instrument was 12 % vs. 1.5 % for culture, with the addition of the instrument’s detection of viruses in 23 samples. Median cTAT for the instrument was 4.2 hours vs. 2 days for culture. Use of the instrument was associated with a statistically significant shorter antibiotic exposure of bacteria-negative vs. positive patients of five days, measured as from the time of reporting of laboratory results to the responsible clinicians. Similarly, its use was associated with a significantly shorter hospitalization for all-negative patients (five days) compared to those with any microorganism detected by it. No statistically significant differences in patient outcome were found due to its use, nor by its detection of any microorganisms. Conclusion: Point-of-care tests provide laboratory means to settings without laboratory capacity and to situations in need of timely results, and we could show how rapid molecular methods for CSF analysis could benefit paediatric children with suspected CNS infection. Yet, without any observed benefits in patient outcome, and at a cost not financially bearable in most lowresource settings. Contextually fit POCTs for paediatric CNS infections are needed in lowresource settings. Yet, there are design flaws in current POCTs and in implementations for their use, limiting their clinical benefits. Collaborative engagement of product developers, clinicians, laboratory professionals, and health policymakers would better serve low-resource settings with contextually fit POCTs and allow for their optimized implementation. The ‘POCTEST' framework for such an engagement is proposed in this thesis. Finally, as we provided proof of concept for a newly developed paper-printed molecular method, we will pursue its development towards contextual clinical utility in low-resource settings. Should we succeed, we hope to contribute to a decrease in preventable childhood mortality in such settings

Bench-to-bedside review: Future novel diagnostics for sepsis - a systems biology approach

The early, accurate diagnosis and risk stratification of sepsis remains an important challenge in the critically ill. Since traditional biomarker strategies have not yielded a gold standard marker for sepsis, focus is shifting towards novel strategies that improve assessment capabilities. The combination of technological advancements and information generated through the human genome project positions systems biology at the forefront of biomarker discovery. While previously available, developments in the technologies focusing on DNA, gene expression, gene regulatory mechanisms, protein and metabolite discovery have made these tools more feasible to implement and less costly, and they have taken on an enhanced capacity such that they are ripe for utilization as tools to advance our knowledge and clinical research. Medicine is in a genome-level era that can leverage the assessment of thousands of molecular signals beyond simply measuring selected circulating proteins. Genomics is the study of the entire complement of genetic material of an individual. Epigenetics is the regulation of gene activity by reversible modifications of the DNA. Transcriptomics is the quantification of the relative levels of messenger RNA for a large number of genes in specific cells or tissues to measure differences in the expression levels of different genes, and the utilization of patterns of differential gene expression to characterize different biological states of a tissue. Proteomics is the large-scale study of proteins. Metabolomics is the study of the small molecule profiles that are the terminal downstream products of the genome and consists of the total complement of all low-molecular-weight molecules that cellular processes leave behind. Taken together, these individual fields of study may be linked during a systems biology approach. There remains a valuable opportunity to deploy these technologies further in human research. The techniques described in this paper not only have the potential to increase the spectrum of diagnostic and prognostic biomarkers in sepsis, but they may also enable the discovery of new disease pathways. This may in turn lead us to improved therapeutic targets. The objective of this paper is to provide an overview and basic framework for clinicians and clinical researchers to better understand the 'omics technologies' to enhance further use of these valuable tools

Human genetics and genomics a decade after the release of the draft sequence of the human genome

10.1186/1479-7364-5-6-577Human Genomics56577-62

Exploring uncertainties regarding unsolicited findings in genetic testing

Objectives: Non-normative uncertainty (uncertainty about empirical facts) and normative uncertainty (uncertainty about moral values or beliefs) regarding unsolicited findings (UFs) might play an important role in clinical genetics. Identifying normative uncertainty is of special interest since it might guide towards novel directions for counseling practice. This study aims to gain insight into the role of non-normative and normative uncertainty regarding UFs, as expressed by counselees and counselors. Methods: We performed a secondary qualitative analysis of interviews with counselees (n = 20) and counselors (n = 20) who had been confronted with UFs. Following a deductive approach, we used Han et al.’s existing theoretical framework of uncertainty, in which we additionally incorporated normative uncertainty. Results: Major issues of non-normative uncertainty were practical and personal for counselees, whilst counselors’ uncertainty pertained mainly to scientific issues. Normative uncertainty was a major theme throughout the interviews. We encountered the moral conflicts of autonomy vs. beneficence and non-maleficence and of autonomy vs. truthfulness. Conclusion: Non-normative uncertainty regarding UFs highlights the need to gain more insight in their penetrance and clinical utility. This study suggests moral conflicts are a major source of feelings of uncertainty in clinical genetics. Practice implications: Exploring counselees’ non-normative uncertainties and normative conflicts seems a prerequisite to optimize genetic counseling.</p